Protocol for Re-using of Qiagen tips

Song-Kun Shyue shsu at HEART.MED.UTH.TMC.EDU
Fri Feb 16 12:04:48 EST 1996


Hi bio-net user:
       
        This is the procotol I send months ago with a little modification,
and I test it again and it just works very well (about 130 ug from each 100
tip). 

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1. For the tip 100, after eluted you DNA wash the column with 2 ml or more
elution buffer to get rid of trace DNA.

2. Wash column with 2 to 3 ml equilibrium buffer, after about 1 ml folw out
stop the flow by wraping with parafilm or cap and seal top with parafilm to
keep column wet that will make it easier to reuse next time. 

3. Kept tip up on room temp. or 4 C until next use. (Next time, just wash
with fresh Equil. buffer then it is read to load the lysate.)

        
        I have tried two cloumns one follow the protocol above the other was
dry after I washed with equil. buffer. the first one gave me about 130 ug
DNA from 10 ml 2xYT culture! 

        One critical step for high yield is to lyse your cells completely
when P2 solution added. If not complete clear after P2 added, add some more
P2 solution. For 15 ml 2YT or 30 ml LB you may need up to 7 ml of P2 to lyse
all cells. Most problem with low yield comes from the incomplete lyse of
cells. I never have the overloaded problem as the company claimed. In
addition, make sure the cells have DNA in it by runing 5 ul of lysate on gel
after P3 (low voltage, about 50 V, get DNA runs into gel slower, then used
high V. to save time) to see if there is DNA in your sample before load into
Tip to save your time and money.

        Good luck to everyone!




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